Potency testing of up to sixteen cannabinoids in hemp-infused edibles using liquid chromatography diode array detector with optional confirmation of identity by electrospray ionization time-of-flight mass spectrometry.

A LC-DAD method for potency testing of up to sixteen cannabinoids has been developed, validated, and applied for analysis of twenty hemp-infused edibles encompassing a broad range of complex matrices. The method was validated according to ISO 17025 guidelines and met requirements. Samples or their uniform water-dispersions were extracted by methanol under homogenization through pulverization and/or ultrasonication. By spiking abnormal cannabidiol, a cannabinoid not naturally present in hemp, into each sample, extraction recovery was tracked in real time, obtaining 90 to 108% in triplicates with relative standard deviations of 0.5 to 6.5%. The linear calibration range was between 0.008 and 10% (w/w) for each cannabinoid using a 250 µg/mL solution of hemp-infused edibles, except for drinks (sparkling water and tea), where it was between 0.0008 and 1% (w/w) using a 2.5 mg/mL solution. ESI/TOFMS confirmed a good method specificity, i.e., without any false positive identification of individual cannabinoid.

A Novel Method Based on Hydrodynamic Cavitation for Improving Nitric Oxide Removal Performance of NaClO2.

In the removal of nitric oxide (NO) by sodium chlorite (NaClO2), the NaClO2 concentration is usually increased, and an alkaline absorbent is added to improve the NO removal efficiency. However, this increases the cost of denitrification. This study is the first to use hydrodynamic cavitation (HC) combined with NaClO2 for wet denitrification. Under optimal experimental conditions, when 3.0 L of NaClO2 with a concentration of 1.00 mmol/L was used to treat NO (concentration: 1000 ppmv and flow rate: 1.0 L/min), 100% of nitrogen oxides (NOx) could be removed in 8.22 min. Furthermore, the NO removal efficiency remained at 100% over the next 6.92 min. Furthermore, the formation of ClO2 by NaClO2 is affected by pH. The initial NOx removal efficiency was 84.8-54.8% for initial pH = 4.00-7.00. The initial NOx removal efficiency increases as the initial pH decreases. When the initial pH was 3.50, the initial NOx removal efficiency reached 100% under the synergistic effect of HC. Therefore, this method enhances the oxidation capacity of NaClO2 through HC, realizes high-efficiency denitrification with low NaClO2 concentration (1.00 mmol/L), and has better practicability for the treatment of NOx from ships.

MicroRNA-489-3p aggravates neuronal apoptosis and oxidative stress after cerebral ischemia-reperfusion injury.

Cerebral ischemia-reperfusion injury (CIRI) mostly occurs in the treatment stage of ischemic diseases and aggravate brain tissue damage. Although studies have demonstrated that miR-489-3p is closely related to CIRI, the effects of miR-489-3p on neural function in CIRI have not been directly studied. The transient middle cerebral artery occlusion (tMCAO) model was established by suture method, and the corresponding plasmids that interfered with the expression of miR-489-3p or Sirtuin1 (SIRT1) were injected into the model mice, and the behavioral changes of the mice were observed. Then the concentration of serum neuronal injury markers and oxidative stress indices were examined. Next, the pathological conditions, neuronal loss and apoptosis of brain tissue were observed by hematoxylin-eosin staining, Nissl staining, and Transferase-mediated deoxyuridine triphosphate-biotin nick end labeling staining. Finally, the hemoglobin content and cerebral edema in the mouse brain were determined. In addition, the expression levels of miR-489-3p and SIRT1 were detected by reverse transcription quantitative polymerase chain reaction or Western blot, and the targeting relationship between miR-489-3p and SIRT1 was verified by bioinformatics analysis and luciferase reporter assay. The experimental results found that in tMCAO mice, miR-489-3p in brain tissue was up-regulated and SIRT1 was down-regulated. Down-regulating miR-489-3p or up-regulating SIRT1 ameliorated behavioral dysfunction, neuronal damage and apoptosis, oxidative stress and brain histopathology. miR-489-3p targeted the regulation of SIRT1 expression, and down-regulating SIRT1 can reverse the protective effect of silenced miR-489-3p on brain injury. Taken together, by targeting SIRT1, elevated miR-489-3p aggravates CIRI-induced neuronal apoptosis and oxidative stress.

Potency testing of up to twenty cannabinoids by liquid chromatography diode array detector with optional electrospray ionization time-of-flight mass spectrometry.

The growing popularity of Cannabis sativa L. and its widespread use for medical and recreational purposes have created an urgent demand of accurate and reliable analytical methods to identify and quantify a growing number of cannabinoids. To meet this demand, a liquid chromatography diode array detector (LC-DAD) method has been developed, validated, and applied in analysis of cannabinoids in nine samples of plant materials of marijuana, six samples of marijuana cigarettes, five samples of hemp flowers, one sample of hemp cigarette, and two samples of Δ8-tetrahydrocannabinol (Δ8-THC) fortified hemp flowers. The method has achieved significant improvements over published methods, which was characterized of 20 targeted cannabinoids, 18 quantified cannabinoids, baseline resolution among quantified cannabinoids, a low limit of quantification (0.02 µg mL-1), a wide linear range (0.02-25 µg mL-1 or 0.04-50% (w/w)), a unique experiment to track the recovery of sample preparation in real time by spiking abnormal cannabidiol (CBD), and a well-assessed specificity by electrospray ionization time-of-flight mass spectrometry (ESI/TOFMS). Precision and accuracy were assessed using quality control (QC) samples at three concentration levels, i.e., 0.02, 0.5, and 12.5 µg mL-1, in triplicates with inter-day and intra-day precision of less than 15% relative standard deviation (RSD) and accuracy of less than ±15% relative error, therefore meeting the requirements by the ISO 17025 standards. Additionally, ESI/TOFMS has discovered seven unknown cannabinoids, including one structural isomer of cannabigerol (CBG), one structural isomer of cannabinolic acid (CBNA), four structural isomers of Δ9-THC, and one structural isomer of Δ9-THC acetate. Furthermore, it uncovered that one of the two samples of Δ8-THC fortified hemp flowers contained 5.16% (w/w) Δ9-THC, an alarm to the current Δ8-THC craze by the public.

Development of a validated method for rapid quantification of up to sixteen cannabinoids using ultra-high-performance liquid chromatography diode-array detector with optional electrospray ionization time-of-flight mass spectrometry detection.

Due to the recent legalization of medical and recreational Cannabis in many countries in the world, there has been an increasing demand for accurate quantification of a growing number of cannabinoids. To meet this challenge, a method for rapid quantification of up to sixteen cannabinoids using ultra-high-performance liquid chromatography diode-array detector (UHPLC-DAD) has been developed, validated and used in the analysis of hemp concentrates. While published LC-UV methods were usually for twelve or less cannabinoids and might not achieve baseline separation of some critical pairs of cannabinoids, e.g., CBG/CBD (cannabigerol/cannabidiol) and Δ9-THC/Δ8-THC (tetrahydrocannabinol), in this study a systematic separation optimization led to a resolution of 1.7 for both pairs. The linear calibration range of all cannabinoids were between 0.02 to 25 µg/mL in methanol, leading to the quantification of 0.1 to 125% (w/w) individual cannabinoids in hemp concentrates after they were mixed with methanol at 20 µg/mL and analyzed after ultrasonication, centrifugation and filtration. The analytical results showed that none of the nine analyzed samples contained any of the seven acidic cannabinoids, but all the nine neutral cannabinoids were detected, with average content ranging from 0.10 to 93.23% (w/w) and RSD (relative standard deviation) values from 0.3 to 11.2% in triplicates. Particularly, two hemp concentrates, i.e., delta 8 hemp distillate and delta 8 hemp shatter contained 9.35 and 11.33% (w/w) Δ9-THC, respectively. While published recovery experiments were limited by the unavailability of cannabinoid-free matrix and the high cost of cannabinoid standards, this problem was solved by spiking abnormal CBD, a cannabinoid not naturally present in Cannabis plants and commercially available with a reasonable price, into the samples. The obtained average recovery ranged from 94.8 to 103.6% with RSD values from 1.5 to 9.0% in triplicates for the nine analyzed samples. Electrospray ionization time-of-flight mass spectrometry (ESI/TOFMS) confirmed the good specificity of the UHPLC-DAD method, i.e., without any false positive identification of individual cannabinoids, and discovered six untargeted cannabinoids that were structural isomers of Δ9-THC in the nine hemp concentrate samples.

Insight into the promoting effect of support pretreatment with sulfate acid on selective catalytic reduction performance of CeO2/ZrO2 catalysts.

In this paper, sulfated ZrO2 were synthesized via precipitation and impregnation method, and the promoting effects of support sulfation on selective catalytic reduction (SCR) performance of CeO2/ZrO2 catalysts were investigated. The results revealed that sulfated ZrO2 could significantly enhance the SCR activity of CeO2/ZrO2 catalysts in a wide temperature range. Especially when S/Zr molar ratio was 0.1, CeO2/ZrO2-0.1S catalyst exhibited a large operating temperature window of 251 âˆ¼ 500 °C and its N2 selectivity was 100 % in the temperature range of 150 âˆ¼ 500 °C. Moreover, CeO2/ZrO2-0.1S catalyst possessed a superior low-temperature activity over 0.1S-CeO2/ZrO2 catalyst. After exposing to 100 ppm SO2 for 15 h, a high NO conversion efficiency of CeO2/ZrO2-0.1S catalyst (90.7 %) could still be reached. The characterization results indicated that ZrO2 treated with a proper dosage of sulfate acid was beneficial to enlarge the specific surface area greatly. Sulfated ZrO2 was also in favor of promoting the transformation of CeO2 from crystalline state to highly-dispersed amorphous state, and inhibiting the transformation of ZrO2 from tetragonal to monoclinic phase. It could also enhance the total surface acidity greatly with an increase in both Brønsted acid sites and Lewis acid sites, thus significantly improving NH3 adsorption on catalyst surface. Besides, the promoting effect of support sulfation on SCR performance of CeO2/ZrO2 catalysts was also related with the enhanced redox property, higher Ce3+/(Ce3++Ce4+) ratio and abundant surface chemisorbed labile oxygen. The in-situ DRIFTS results implied that nitrate species coordinated on the surface of CeO2/ZrO2-0.1S catalyst could participate in the Selective catalytic reduction with ammonia (NH3-SCR) reactions at either medium or high temperature, suggesting that both Eley-Rideal (E-R) and Langmuir-Hinshelwood (L-H) mechanisms might be followed in SCR reactions.

Nitrogen direct analysis in real time time-of-flight mass spectrometry (N2 DART-TOFMS) for rapid screening of forensic drugs.

RATIONALE: Over the last ten years, helium direct analysis in real time time-of-flight mass spectrometry (He DART-TOFMS) has become an established technique in rapid screening of forensic drugs to decrease the time necessary to triage forensic drug cases, therefore contributing to backlog reduction and more timely criminal prosecution. Recently, we demonstrated that N2 DART was able to efficiently ionize all polar compounds except for a few extremely small ones such as methanol and acetonitrile. Therefore, N2 DART-TOFMS should be a suitable technique for rapid screening of forensic drugs. METHODS: Nitrogen direct analysis in real time time-of-flight mass spectrometry (N2 DART-TOFMS) was performed using a JEOL AccuTOF mass spectrometer with an IonSense DART-100 ion source. A 3-min analytical protocol was used for the analysis of each sample. Sample introduction was accomplished by moving the closed end of a melting point capillary where approximately 1 µL sample solution was deposited or the exposed inside of a freshly cut tablet across the N2 gas stream between the DART-100 ion source and orifice 1 of the AccuTOF. RESULTS: Ten commonly abused drugs, eight synthetic cannabinoids and four controlled prescription drugs (CPDs) were analyzed. The limit of detection (LOD) was determined to be approximately 10 µg/mL or 10 pg in quantities. All drugs at the LOD level were positively identified using their [M + H]+ ions with mass errors less than 5 mDa. The identification were further supported by in-source fragment ions and characteristic N2 DART ions that are not commonly generated by He DART, e.g. [M + H + O]+ and [M + H + 2O]+ ions. CONCLUSIONS: It was concluded that the 3-min analytical protocol could be utilized in the analysis of seized drugs in the form of tablets and powders or prepared in solution. In consideration that N2 is readily available in the air and He is a non-renewable resource, N2 DART-TOFMS is a greener, cheaper and more convenient alternative to He DART-TOFMS in rapid screening of forensic drugs.

Leguminous supplementation increases the resilience of soil microbial community and nutrients in Chinese fir plantations.

Understory vegetation plays a vital role in the flow of materials and nutrient cycling in plantation ecosystems. Introducing functional plants (one species or a group of plants that share similar characteristics and can play a similar role in an ecological environment) can quickly improve the environment of the soil of a plantation with a single-stand structure suffering from soil degradation. Five stands composed of Chinese fir plants of different ages (young, immature, near-mature, mature, and over-mature stand forests) were supplemented with leguminous plants to determine the effects on soil nutrients and microbial communities. We supplemented the five stands with five different combinations of four non-native plant species, Dalbergia balansae, Taxus chinensis, Spatholobus suberectus, and Kaempferia galangal, as treatments. After one year, plant growth was estimated, and soil samples were collected for laboratory experiments and high-throughput sequencing. Our results show that supplementing the stands with plants increased the nutrient content of the soil and promoted the growth and diversity of soil microbial communities in Chinese fir plantations. Furthermore, the effects of plant supplementation varied according to the age of the stand in the plantation; thus, the positive effects were stronger for young, immature, and near-mature stand forests than they were for mature and over-mature stand forests. Measurements of the microbial diversity in the soil revealed that supplementation increased diversity in the fungal community more than that in the bacterial community. A principal component analysis (PCA) of the five treatments and controls under different forest stands ages demonstrated that microbial communities differed significantly between treatments and controls and that supplementing Chinese fir plantations with leguminous plants had a greater influence on microbial communities than other plants did. Our study suggests that certain leguminous plants can increase soil nutrients and the diversity of soil microbial communities in one year.

Ionization Mechanism of Positive-Ion Nitrogen Direct Analysis in Real Time.

Nitrogen can be an inexpensive alternative to helium used by direct analysis in real time (DART), especially in consideration of the looming helium shortage. Therefore, the ionization mechanism of positive-ion N2 DART has been systematically investigated. Our experiments suggest that a range of metastable nitrogen species with a variety of internal energies existed and all of them were less energetic than metastable helium atoms. However, compounds with ionization energies (IE) equal to or lower than 10.2 eV (all organic compounds except the extremely small ones) can be efficiently ionized. Because N2 DART was unable to efficiently ionize ambient moisture and common organic solvents such as methanol and acetonitrile, the most important ionization mechanism was direct Penning ionization followed by self-protonation of polar compounds generating [M+H]+ ions. On the other hand, N2 DART was able to efficiently ionize ammonia, which was beneficial in the ionization of hydrogen-bonding compounds with proton affinities (PA) weaker than ammonia generating [M+NH4]+ ions and large PAHs generating [M+H]+ ions through proton transfer. N2 DART was also able to efficiently ionize NO, which led to the ionization of nonpolar compounds such as alkanes and small aromatics generating [M-(2m+1)H]+ (m=0,1…) ions. Lastly, metastable nitrogen species was also able to produce oxygen atoms, which resulted in increased oxygen adducts as the polarity of organic compounds decreased. In comparison with He DART, N2 DART was approximately one order of magnitude less sensitive in generating [M+H]+ ions, but could be more sensitive in generating [M+NH4]+ ions. Graphical Abstract ᅟ.

Quantitative real-time monitoring of chemical reactions by autosampling flow injection analysis coupled with atmospheric pressure chemical ionization mass spectrometry.

Although qualitative and/or semiquantitative real-time monitoring of chemical reactions have been reported with a few mass spectrometric approaches, to our knowledge, no quantitative mass spectrometric approach has been reported so far to have a calibration valid up to molar concentrations as required by process control. This is mostly due to the absence of a practical solution that could well address the sample overloading issue. In this study, a novel autosampling flow injection analysis coupled with an atmospheric pressure chemical ionization mass spectrometry (FIA/APCI-MS) system, consisting of a 1 µL automatic internal sample injector, a postinjection splitter with 1:10 splitting ratio, and a detached APCI source connected to the mass spectrometer using a 4.5 in. long, 0.042 in. inner diameter (ID) stainless-steel capillary, was thus introduced. Using this system together with an optional FIA solvent modifier, e.g., 0.05% (v/v) isopropylamine, a linear quantitative calibration up to molar concentration has been achieved with 3.4-7.2% relative standard deviations (RSDs) for 4 replicates. As a result, quantitative real-time monitoring of a model reaction was successfully performed at the 1.63 M level. It is expected that this novel autosampling FIA/APCI-MS system can be used in quantitative real-time monitoring of a wide range of reactions under diverse reaction conditions.

Differentiation of underivatized monosaccharides by atmospheric pressure chemical ionization quadrupole time-of-flight mass spectrometry.

RATIONALE: Differentiation of underivatized monosaccharides is essential in the structural elucidation of oligosaccharides which are closely involved in many life processes. So far, such differentiation has been usually achieved by electrospray ionization mass spectrometry (ESI-MS). As an alternative to ESI-MS, atmospheric pressure chemical ionization mass spectrometry (APCI-MS) should provide complementary results. METHODS: A quadrupole time-of-flight (QTOF) mass spectrometer with accurate mass measurement ability was used with an APCI heated nebulizer ion source because we believe that a recently published article using a single quadrupole mass spectrometer assigned incorrect identities for APCI ions from hexoses. Using APCI-QTOF, the MS(2) and pseudo-MS(3) mass spectra of 11 underivatized monosaccharides were obtained under various collision voltages. The mass spectra were carefully interpreted after accurate mass measurement. RESULTS: Differentiation of three hexoses was achieved by different MS(2) spectra of their [M + NH(4)](+) and [M - H](-) ions. The MS(2) spectra of the [M + NH(4)](+) ions were also used to distinguish methyl α-D-glucose and methyl ß-D-glucose, while the pseudo-MS(3) spectra of the [M + H](+) ions were utilized to differentiate the three hexosamine and N-acetylhexosamine stereoisomers. Unique [M + O(2)](-) ions were observed and their distinctive fragmentation patterns were utilized to differentiate the three hexosamine stereoisomers. CONCLUSIONS: Although ESI coupled with single or triple quadrupole and ion trap mass spectrometers has been widely utilized in the differentiation of monosaccharides, this report demonstrated that APCI-QTOF-MS had its own advantages in achieving the same goal.

Evaluation of direct analysis in real time mass spectrometry for onsite monitoring of batch slurry reactions.

Batch slurry reactions are widely used in the industrial manufacturing of chemicals, pharmaceuticals, petrochemicals and polymers. However, onsite monitoring of batch slurry reactions is still not feasible in production plants due to the challenge in analyzing heterogeneous samples without complicated sample preparation procedures. In this study, direct analysis in real time mass spectrometry (DART-MS) has been evaluated for the onsite monitoring of a model batch slurry reaction. The results suggested that automation of the sampling process of DART-MS is important to achieve quantitative results. With a sampling technique of manual sample deposition on melting point capillaries followed by automatic sample introduction across the helium beam, relative standard deviation (RSD) of the protonated molecule signals from the reaction product of the model batch slurry reaction ranged from 6 to 30%. This RSD range is improved greatly over a sampling technique of manual sample deposition followed by manual sample introduction where the RSDs are up to 110%. Furthermore, with the semi-automated sampling approach, semi-quantitative analysis of slurry samples has been achieved. Better quantification is expected with a fully automated sampling approach.

Ionization mechanism of positive-ion direct analysis in real time: a transient microenvironment concept.

A transient microenvironment mechanism (TMEM) is proposed to address matrix effects for direct analysis in real time (DART). When the DART gas stream is in contact with the sample, a transient microenvironment (TME), which can shield analytes from direct ionization, may be generated through the desorption of the matrix containing the analyte. The DART gas stream can directly ionize the matrix molecules, but the analytes will be ionized primarily through gas-phase ion/molecule reactions with the matrix ions. Experimental results showed that as little as 10 nL of liquid or 10 microg of solid was able to generate an efficient TME. Generated TMEs were able to control the ionization of an analyte below an analyte-to-matrix ratio that was dependent on the DART temperature and the boiling points of the analyte and matrix. TMEs generated by common solvents were studied in detail. The ionization of both polar and nonpolar compounds, present in a solvent or another analyte below a ratio of 1:100, were found to be mainly controlled by the generated TMEs at a DART temperature of 300 degrees C.

Ionization mechanism of negative ion-direct analysis in real time: a comparative study with negative ion-atmospheric pressure photoionization.

The ionization mechanism of negative ion-direct analysis in real time (NI-DART) has been investigated using over 42 compounds, including fullerenes, perfluorocarbons (PFC), organic explosives, phenols, pentafluorobenzyl (PFB) derivatized phenols, anilines, and carboxylic acids, which were previously studied by negative ion-atmospheric pressure photoionization (NI-APPI). NI-DART generated ionization products similar to NI-APPI, which led to four ionization mechanisms, including electron capture (EC), dissociative EC, proton transfer, and anion attachment. These four ionization mechanisms make both NI-DART and NI-APPI capable of ionizing a wider range of compounds than negative ion-atmospheric pressure chemical ionization (APCI) or negative ion-electrospray ionization (ESI). As the operation of NI-DART is much easier than that of NI-APPI and the gas-phase ion chemistry of NI-DART is more easily manipulated than that of NI-APPI, NI-DART can be therefore used to study in detail the ionization mechanism of LC/NI-APPI-MS, which would be a powerful methodology for the quantification of low-polarity compounds. Herein, one such application has been further demonstrated in the detection and identification of background ions from LC solvents and APPI dopants, including water, acetonitrile, chloroform, methylene chloride, methanol, 2-propanol, hexanes, heptane, cyclohexane, acetone, tetrahydrofuran (THF), 1,4-dioxane, toluene, and anisole. Possible reaction pathways leading to the formation of these background ions were further inferred. One of the conclusions from these experiments is that THF and 1,4-dioxane are inappropriate to be used as solvents and/or dopants for LC/NI-APPI-MS due to their high reactivity with source basic ions, leading to many reactant ions in the background.

Liquid chromatography/negative ion atmospheric pressure photoionization mass spectrometry: a highly sensitive method for the analysis of organic explosives.

Gas chromatography/mass spectrometry (GC/MS) is applied to the analysis of volatile and thermally stable compounds, while liquid chromatography/atmospheric pressure chemical ionization mass spectrometry (LC/APCI-MS) and liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) are preferred for the analysis of compounds with solution acid-base chemistry. Because organic explosives are compounds with low polarity and some of them are thermally labile, they have not been very well analyzed by GC/MS, LC/APCI-MS and LC/ESI-MS. Herein, we demonstrate liquid chromatography/negative ion atmospheric pressure photoionization mass spectrometry (LC/NI-APPI-MS) as a novel and highly sensitive method for their analysis. Using LC/NI-APPI-MS, limits of quantification (LOQs) of nitroaromatics and nitramines down to the middle pg range have been achieved in full MS scan mode, which are approximately one order to two orders magnitude lower than those previously reported using GC/MS or LC/APCI-MS. The calibration dynamic ranges achieved by LC/NI-APPI-MS are also wider than those using GC/MS and LC/APCI-MS. The reproducibility of LC/NI-APPI-MS is also very reliable, with the intraday and interday variabilities by coefficient of variation (CV) of 0.2-3.4% and 0.6-1.9% for 2,4,6-trinitrotoluene (2,4,6-TNT).

Evaluation of ceftazidime contents in antibiotic discs by capillary electrophoresis.

Good quality of antibiotic discs is a fundamental prerequisite to accurate antibiotic susceptibility tests. Capillary electrophoresis (CE) is a widely used method for quantitative analysis. Here, using ceftazidime as an example, we report an easy-to-perform strategy to determine ceftazidime content in discs. First, a serial of ceftazidime standard solutions was prepared to determine the detection linearity. Utilizing the background buffer containing 50 mmol/L Na2HPO4, 0.045 mmol/L beta-cyclodextrin and 3.15 mmol/L Tris (hydroxymethyl) aminomethane, ceftazidime of concentrations ranging from 1.875 to 60 microg/ml showed a linear response to detected peak areas. Subsequently, four kinds of ceftazidime discs were selected from different manufacturers. The discs were homogenized using 1 ml deionized water, and then detected by CE after filtration. The results showed that one kind of discs were of poor quality as further confirmed by disc diffusion tests using standard strains. This study proved the potential of CE as an easy choice to perform disc quality control under appropriate conditions.

[Determination of alachlor residue in agricultural products].

A method of the determination of alachlor residue in agricultural products was developed. Four different kinds of objects, corn, peanut, spinach, and orange were selected as representatives of main agricultural products. The sample was extracted with acetone-water (8:2, v/v). The extract was partitioned with dichloromethane, cleaned up by gel permeation chromatography (GPC) and solid phase extraction (SPE) in order to completely separate colorants from alachlor in the sample. The eluate was evaporated under vacuum to nearly dry and diluted with acetone to a definite volume. The solution was determined and identified by gas chromatography-mass spectrometry (GC-MS) using external standard method. The recoveries were 86.0%-98.2%, and relative standard deviations were 5.1%-6.7% with spiked at 0.010-0.200 mg/kg of alachlor in blank samples. The limit of quantification of this method was 0.010 mg/kg.

Negative ion-atmospheric pressure photoionization: electron capture, dissociative electron capture, proton transfer, and anion attachment.

To better guide the development of liquid chromatography/electron capture-atmospheric pressure photoionization-mass spectrometry (LC/EC-APPI-MS) in analysis of low polarity compounds, the ionization mechanism of 19 compounds was studied using dopant assisted negative ion-APPI. Four ionization mechanisms, i.e., EC, dissociative EC, proton transfer, and anion attachment, were identified as being responsible for the ionization of the studied compounds. The mechanisms were found to sometimes compete with each other, resulting in multiple ionization products from the same molecule. However, dissociative EC and proton transfer could also combine to generate the same [M - H](-) ions. Experimental evidence suggests that O(2)(-*), which was directly observed in the APPI source, plays a key role in the formation of [M - H](-) ions by way of proton transfer. Introduction of anions more basic than O(2)(-*), i.e., C(6)H(5)CH(2)(-), into the APPI source, via addition of di-tert-butyl peroxide in the solvent and/or dopant, i.e., toluene, enhanced the deprotonation ability of negative ion-APPI. Although the use of halogenated solvents could hinder efficient EC, dissociative EC, and proton transfer of negative ion-APPI due to their EC ability, the subsequently generated halide anions promoted halide attachment to compounds that otherwise could not be efficiently ionized. With the four available ionization mechanisms, it becomes obvious that negative ion-APPI is capable of ionizing a wider range of compounds than negative ion chemical ionization (NICI), negative ion-atmospheric pressure chemical ionization (negative ion-APCI) or negative ion-electrospray ionization (negative ion-ESI).

Electron capture atmospheric pressure photoionization mass spectrometry: analysis of fullerenes, perfluorinated compounds, and pentafluorobenzyl derivatives.

An electron capture (EC) ionization mechanism has been found to be highly efficient in negative-ion atmospheric pressure photoionization (APPI) for the analysis of compounds with positive electron affinity (EA). Using negative-ion APPI, we first report the sensitive detection of natural electrophores with limited polarity, such as fullerenes and perfluorinated compounds, by mass spectrometry (MS). Using direct infusion on a quadrupole time-of-flight (QTOF) mass spectrometer, the limits of detection (LODs) for C(60) and perfluoromethylcyclohexane were determined to be 0.15 pg (0.2 fmol) and 1 femtoliter (fL) ( approximately 1.5 pg or 4.3 fmol), respectively. As the EA of the analyte increases, the detection sensitivity is enhanced. Making use of the accurate mass measurement capability of the QTOF mass spectrometer, we were able to investigate the elemental composition of the ions in each spectrum and attribute the observed high sensitivity to an EC-initiated ionization process. The proposed EC ionization mechanism is further supported by the observation of a dissociative EC reaction of pentafluorobenzyl (PFB)-derivatized phenols. The analysis of phenols by EC-APPI of their PFB derivatives resulted in very high sensitivity, with the lowest reported LOD of approximately 0.17 pg (0.5 fmol) being for 2,4-dinitrophenol. For future LC/EC-APPI-MS applications, the effect of additives and solvents on sensitivity was also tested and reported.

The principal urinary metabolites of dietary isothiocyanates, N-acetylcysteine conjugates, elicit the same anti-proliferative response as their parent compounds in human bladder cancer cells.

Isothiocyanates (ITCs) are a class of well-known cancerpreventive phytochemicals, but are primarily disposed of and concentrated in the urine as N-acetylcysteine conjugates (NAC-ITCs) in vivo. Because human urinary bladder cancers occur almost exclusively in the bladder epithelium, which is directly exposed to the urine stored in the bladder, we undertook to examine the anti-cancer activity of NAC-ITCs in cultured human bladder cancer cells. In this paper, we report that the NAC conjugates of four naturally occurring ITCs, including allyl ITC, benzyl ITC (BITC), phenethyl ITC and sulforaphane, potently inhibited the growth of cells derived from both low-grade superficial and high-grade invasive human bladder cancers and drug-resistant bladder cancer cells. Moreover, the growth-inhibitory potencies were similar between the conjugates and their parent compounds. Further study of NAC-BITC and BITC as model compounds showed that both compounds accumulated in cells predominantly as the glutathione conjugate of BITC, but the accumulation of the former was slower. Moreover, both compounds also demonstrated the same anti-proliferative mechanisms: causing the cleavage of the same set of caspases (caspase-3, -8 and -9) in apoptosis induction, arresting cells in the same phases (S and G2/M) and targeting the same cell cycle regulator (Cdc25C), although a longer treatment time or slightly higher doses were needed for NAC-BITC to achieve the same effect as BITC, presumably due to slower cellular uptake of NAC-BITC. These data show that the NAC-ITCs are biologically similar to their parent compounds and are highly effective against human bladder cancer cells.